We propose to sequence selected regions of tryptophan (trp) operon messenger RNA from E. coli and other Enterobacteria, and determine whether these regions exhibit secondary structure. The trp mRNA regions we will study include: 1) The translation initiation and termination sequences of trpA mRNA, the messenger region for the tryptophan synthetase alpha chain, 2) the sequence specifying the initial 100 amino acids of the alpha chain, 3) the post-amino acid specifying region of the messenger, and 4) the translation punctuation sequence between trpB mRNA and trpA mRNA. We will also determine whether synonymous codon changes have accumulated in trpA mRNA during the course of evolution and whether the trpB mRNA-trpA mRNA punctuation region and the post-amino acid specifying region of the messenger have been more highly conserved than the amino acid sequence spcifying portion. We will continue our analyses of intergeneric intragenic recombination between trpA of E. coli and trpA of Salmonella typhimurium. We will determine the fraction of such recombinants which have defective alpha chains. Representative alpha chains will be isolated and their amino acid sequences compared with those of the parental 2 chains. An attempt will be made to assess the enzymatic capabilities of hybrid alpha chains. Mutants will be sought in which the frequency of intergeneric intragenic recombination is increased appreciably. We will isolate and characterize a new class of polarity-relief suppressors. An in vitro coupled transcription-translation system will be used in the detection of proteins involved in mRNA degradation and polarity.